The present invention relates to instruments and to methods of measuring low levels of consumption and production of certain components, typically oxygen and carbon dioxide, in a fluid medium, such as air, by plant and animal tissues, cell cultures, synthetic resins and metals. More particularly, the present invention relates to such instruments and methods which compensate for interfering factors, such as sensor drift and pressure/temperature fluctuations.
The measurement of oxygen consumption and carbon dioxide production is applied in several fields. In entomology, such measurements are helpful in determining insect food sources. In the perishable food industry, respiration measurements are used to evaluate means of retarding ripening. Another application is in measuring the biological activity of bacterial cultures, sludge, and other hazardous organic materials. Respirometers are also used to measure oxidation of metals and oxygen uptake by synthetic resins during polymerization.
Various methods and instruments have been used in the past to measure low levels of respiratory gas exchange. Oxygen consumption is commonly determined by measuring changes in the volume of gas in a chamber as oxygen is consumed by the tissue or other sample. The Warburg apparatus, a relatively well known example of such a volumetric device, employs a solution of potassium hydroxide in the sample chamber to absorb carbon dioxide produced by the sample. The change in the volume of the air in the sample chamber is measured by a manometric tube which is connected to the sample chamber on one end and is open to the atmosphere on the other end. Oxygen consumption is assumed to be the sole source of air volume change within the sample chamber. This assumption, however, is unreliable because other factors, such as the temperature of the sample chamber and the barometric pressure, can change its air volume. Such temperature-induced air volume changes can be minimized by placing the apparatus in a temperature-controlled water bath, but even the best water baths cannot regulate the temperature precisely enough for the present low-level respiration applications.
Differential volumetric respirometers, such as the apparatus disclosed in U.S. Pat. No. 3,313,157 to Gilson, seek to improve upon the Warburg apparatus. The adverse effects of temperature changes are offset by employing a reference chamber substantially equal in volume to the sample chamber, under the principal that any temperature change affects the volumes of the gases in the two chambers equally. Oxygen consumption is measured by the difference between the volume changes in the two chambers. However, the assumption that the temperatures of the chambers are the same is unreliable if the sample generates it own heat, as is likely in oxidative reactions.
The Warburg and differential volumetric respirometers also lack the capacity to easily measure carbon dioxide production. Such knowledge is important in many metabolic measurements, since the carbon dioxide production rate is used together with the oxygen consumption rate to compute a respiratory quotient which indicates what foodstuff is being metabolized. In the Warburg device, carbon dioxide production can be determined only by chemical analysis of the potassium hydroxide solution which presumably absorbs the entire output of this gas. Accurate chemical analysis is difficult and time consuming, and only a single determination can be made at the end of the study. Also, in plants, carbon dioxide is consumed, rather than produced, and carbon dioxide consumption cannot be measured using this technique.
Closed respirometers which employ gas analyzers to measure consumption and production of specific gases are also known in the art. Gas analyzers, however, often fluctuate or drift as a result of environmental influences, such as pressure and temperature changes. Typically, drift in the gas analyzer is corrected by manual recalibration of the sensor using a known reference gas prior to each sample gas measurement. However, U.K. Patent No. 2049192A to Austin et al. discloses a differential-type carbon dioxide analyzer in which an infrared beam is alternately directed through a first cell containing a sample gas and a second cell containing a reference gas. The Austin et al. device produces an output signal proportional to the difference between the signals produced after passage of the two beams through the two analyzer cells. To a degree, the Austin et al. respirometer corrects for drift in the gas analyzers, but it has limited applicability because the infrared absorption device it employs cannot detect many other gases, such as oxygen. Furthermore, the Austin et al. device does not fully compensate for sample-induced sensor drift. For example, gas pressure and content variations in the sample cell caused by water vapor from the sample would not be found in the reference cell. Hence, the output signal would not compensate for such drift.
Thus, the present inventors were faced with the problems of accurately measuring changes in the composition of a multi-component fluid medium resulting from low levels of respiration or oxidation while accounting for variables, such as temperature and pressure changes, within the system.